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It has been suggested to longitudinally monitor Insulin-like growth factor I (IGF-I) as a biomarker for the detection of recombinant growth hormone (GH). Subsequently, it is of interest to understand any confounders of endogenous IGF-I. Herein we have studied if serum IGF-I concentration is affected by the intake of anabolic androgenic steroids (AAS) and the potential connection between IGF-I and klotho protein. Moreover, the usefulness of klotho as a biomarker for recombinant GH intake was assessed in healthy male volunteers. An ongoing administration of AAS did not affect the levels of IGF-I. Klotho protein was ~30% higher in men with an ongoing AAS use compared to those with previous (2 months ago) AAS use, and the serum klotho protein correlated negatively with luteinizing hormone (LH) (

The detection of low doses of recombinant growth hormone is a challenge in antidoping testing. Future testing may lead toward the longitudinal monitoring of IGF-I and P-III-NP in an endocrine module. Additional biomarkers, for example vitamin D binding protein, alpha-HS-glycoprotein, fibronectin 1, and decorin have been identified in different omics studies. This was a longitudinal study of the usefulness of these putative biomarkers in relation to 2 weeks administration of low doses of recombinant growth hormone in healthy male volunteers. Moreover, the hematological parameters included in the athlete biological passport were studied as well as the serum concentration of testosterone and dihydrotestosterone. Fibronectin 1 increased by 20% during the treatment period (P ˂ 0.05), confirming the previous finding. Alpha-HS-glycoprotein decreased by 25% up to 3 weeks after treatment (P ˂ 0.05), contradicting previous results. The addition of fibronectin 1 increased the likelihood of detecting recombinant growth hormone intake based on individual calculated thresholds in some of the participants compared with the GH2000, IGF-I, and P-III-NP. The multiplication of fibronectin 1 concentration by IGF-I resulted in the most profound (up to 4-fold) changes. A minor 15% increase (P = 0.003) in the reticulocyte percentage was observed, but the changes did not lead to any atypical profile based on individual passport thresholds. Vitamin D binding protein, decorin, testosterone, and dihydrotestosterone were not affected by growth hormone. Dihydrotestosterone sulfate was negatively correlated with IGF-I at baseline (R = -0.50, P = 0.003) and post dose (R = -0.59, P = 0.01). In conclusion, fibronectin 1 was verified as a promising future biomarker for detecting low doses of recombinant growth hormone.

The efficiency of the immunopurification step of urinary erythropoietin (EPO) and recombinant forms is important for their optimal detection in antidoping screening. Previous investigations of immunopurification techniques have been done for immunomagnetic beads, EPO Purification Kit (EPK) columns (MAIIA Diagnostics), and enzyme-linked immunosorbent assay (ELISA) microplates (Stemcell Technologies) conjugated/coated with anti-EPO antibodies. In this study, a new immunopurification technique using anti-EPO sepharose gel beads, developed by MAIIA Diagnostics, to simplify and minimize sample handling was evaluated. This EPO Purification Gel Kit (EPGK) was compared with our current routine EPK for limit of detection (LOD). Linearity, recovery, repeatability, sample incubation time, and sample volume were also evaluated for EPGK. The LODs and linearity for EPK and EPGK were comparable to each other and the recovery for BRP, NESP, CERA, and EPO-Fc were within the range of other studies, and concentration of the sample eluate improved the recovery results. Little variation was seen within days, between days, and between operators. A 90 minute incubation of the sample with the sepharose gel beads is sufficient for most of the erythropoiesis stimulating agents (ESAs) tested, with 10 mL being an optimal sample volume for EPGK. The improved sample handling, higher sample throughput and the reduced working time demonstrate that the EPGK is a better alternative to the current MAIIA EPK immunopurification method for urine. The EPO Purification Gel Kit (from MAIIA Diagnostics) was evaluated and validated for immunopurification of endogenous erythropoietin and exogenous erythropoiesis stimulating agents from urine samples. The kit was a better alternative to that currently used (EPO Purification Kit) in many antidoping laboratories because it improves sample handling and increases sample throughput.

To detect doping with growth hormone (GH), GH isoform and biomarkers tests are available. Both methods use population-based decision limits. Future testing in anti-doping is progressing toward individual-based reference ranges, and it is possible that with such an approach the sensitivity to detect GH doping may increase. In addition to monitoring different proteins, the use of miRNAs as future GH biomarkers has been discussed. Here we have longitudinally studied the serum concentrations of IGF-I, P-III-NP and the different GH isoforms in nine healthy men prior to, during and after two weeks' administration with low doses (1 and 4 IU/day) of recGH. Moreover, three putative miRNAs were analyzed. The results show that 80% of the participants were identified as atypical findings using the GH isoform test. However, the participants were only positive 1.5-3 hours directly after an injection. Only one of the participants reached a GH-2000 score indicative of doping when a population-based decision limit was applied. When IGF-I and P-III-NP were longitudinally monitored, 88% of the participants were identified above an individual upper threshold arbitrarily calculated as three standard deviations above the mean values of four baseline samples. The miRNA levels displayed large intra-subject variations that did not change in relation to recGH administration. Our results show that the GH isoform test is very sensitive in detecting low doses of recGH but with a short detection window. Moreover, longitudinally monitoring of IGF-I and P-III-NP may be a promising future approach to detect GH doping.

Cancer cells that escape induction therapy are a major cause of relapse. Understanding metabolic alterations associated with drug resistance opens up unexplored opportunities for the development of new therapeutic strategies. Here, we applied a broad spectrum of technologies including RNA sequencing, global untargeted metabolomics, and stable isotope labeling mass spectrometry to identify metabolic changes in P-glycoprotein overexpressing T-cell acute lymphoblastic leukemia (ALL) cells, which escaped a therapeutically relevant daunorubicin treatment. We show that compared with sensitive ALL cells, resistant leukemia cells possess a fundamentally rewired central metabolism characterized by reduced dependence on glutamine despite a lack of expression of glutamate-ammonia ligase (GLUL), a higher demand for glucose and an altered rate of fatty acid β-oxidation, accompanied by a decreased pantothenic acid uptake capacity. We experimentally validate our findings by selectively targeting components of this metabolic switch, using approved drugs and starvation approaches followed by cell viability analyses in both the ALL cells and in an acute myeloid leukemia (AML) sensitive/resistant cell line pair. We demonstrate how comparative metabolomics and RNA expression profiling of drug-sensitive and -resistant cells expose targetable metabolic changes and potential resistance markers. Our results show that drug resistance is associated with significant metabolic costs in cancer cells, which could be exploited using new therapeutic strategies.

Random graphs (or networks) have gained a significant increase of interest due to its popularity in modeling and simulating many complex real-world systems. Degree sequence is one of the most important aspects of these systems. Random graphs with a given degree sequence can capture many characteristics like dependent edges and non-binomial degree distribution that are absent in many classical random graph models such as the Erd\H{o}s-R\'{e}nyi graph model. In addition, they have important applications in the uniform sampling of random graphs, counting the number of graphs having the same degree sequence, as well as in string theory, random matrix theory, and matching theory. In this paper, we present an OpenMP-based shared-memory parallel algorithm for generating a random graph with a prescribed degree sequence, which achieves a speedup of 20.5 with 32 cores. One of the steps in our parallel algorithm requires checking the Erd\H{o}s-Gallai characterization, i.e., whether there exists a graph obeying the given degree sequence, in parallel. This paper presents the first non-trivial parallel algorithm for checking the Erd\H{o}s-Gallai characterization, which achieves a speedup of 23 using 32 cores., Comment: 10 pages

Leukemic cells isolated from most patients with acute myelogenous leukemia (AML) have higher low density lipoprotein (LDL) uptake than normal mononuclear blood cells. Little is known, however, about the mechanism behind the elevated LDL uptake. We investigated if AML cells secrete factors that stimulate cellular LDL uptake. Mononuclear blood cells were isolated from peripheral blood from 42 patients with AML at diagnosis. Cellular LDL uptake was determined from the degradation rate of 125I-labelled LDL. Conditioned media from AML cells stimulated the LDL degradation in the leukemic cell lines KG1 and HL60, and in isolated AML cells. The stimulatory effect correlated with the LDL degradation in the AML cells directly after isolation from blood. Conditioned media also autostimulated LDL degradation in the AML cells themselves. Concentrations of IL-6 and IL-8 in AML cell conditioned media correlated with the LDL degradation in AML cells directly after isolation from blood. Addition of R-TNF-α, but not IL-6 or IL-8, stimulated LDL degradation in HL60, KG1, and AML cells. The LDL degradation in AML cells could be inhibited by a LDL receptor blocking antibody. AML cells secrete factors that stimulate LDL uptake in a paracrine and autocrine pattern which open up therapeutic possibilities to inhibit the uptake of LDL by administration of antibodies to these factors.

The cholesterol metabolism is essential for cancer cell proliferation. We found the expression of genes involved in the cholesterol biosynthesis pathway up-regulated in the daunorubicin-resistant leukemia cell line CEM/R2, which is a daughter cell line to the leukemia cell line CCRF-CEM (CEM). Cellular 2H2O labelling, mass spectrometry, and isotopomer analysis revealed an increase in lanosterol synthesis which was not accompanied by an increase in cholesterol flux or pool size in CEM/R2 cells. Exogenous addition of lanosterol had a negative effect on CEM/R2 and a positive effect on sensitive CEM cell viability. Treatment of CEM and CEM/R2 cells with cholesterol biosynthesis inhibitors acting on the enzymes squalene epoxidase and lanosterol synthase, both also involved in the 24,25-epoxycholesterol shunt pathway, revealed a connection of this pathway to lanosterol turnover. Our data highlight that an increased lanosterol flux poses a metabolic weakness of resistant cells that potentially could be therapeutically exploited. Electronic supplementary material The online version of this article (doi:10.1007/s12032-015-0717-5) contains supplementary material, which is available to authorized users.